The structure required for an inhibitor against a protease was studied by protein engineering with the use of protease inhibitor proteins produced by bacteria, mainly Streptomyces subtilisin inhibitor (SSI).The center of reactive site of SSI (methionine) was replaced by other 19 kinds of amino acid. Mutants replaced by a basic amino acid (lysine or arginine) tended to inhibit trypsin that splits a protein at the positions of basic amino acid residues, although the wild type SSI inhibits trypsin scarcely. Only the lysine mutant inhibited lysyl endopeptidase. The mutant SSIs replaced by an aromatic amino acid (tryptophane, tyrosine, and phenylalanine) tended to inhibit chymotrypsin, which splits a protein at the aromatic amino acid residues. Single amino acid change caused alteration of the functional specificity of a protein. Thus, the reactive site in a protease inhibitor should be similar to a substrate structure to fit to the active center of a protease.The reactive site is supported by other parts of SSI molecule. For example, if the S-S bridge near the reactive site was taken off by replacing two cysteine residues by serine residues, SSI tended to a substrate for proteases. From these experiments we reached to the conclusion that the reactive site in an inhibitor protein molecule should be supported firmly.The study of physical properties of the similar protease inhibitors, which are obtained not only from bacteria but also fungi and plant seeds, and of the artificially modified inhibitors was carried out. The interaction of these inhibitors and a protease was also studied by X-ray analysis, NMR, and other spectral analysis. From these results the following conclusion was obtained: A fine structural change is caused easily in the reactive site and the part near C-terminal, so that the reactive site causes induced fitting on the interaction with the active site of a protease.